It is known that many pathologic conditions cause changes in the .alpha.-amylase levels in body fluids such as serum, urine, etc., and diagnostic methods for measuring the .alpha.-amylase concentration in body fluids have been developed. These methods are usually based on the degrading effect of .alpha.-amylase on starch products, the enzyme activity being determined by measuring--directly or indirectly--the degree of degradation of a starch-like substrate. Examples of such known methods are the so-called "blue starch method" and the so-called "saccharogenic method". In the first mentioned method the substrate is insoluble, cross-linked starch, whereas soluable (Zulkowsky) starch is used in the saccharogenic method. These and other methods for measuring the .alpha.-amylase concentration in body fluids are well known in the art. See e.g. O'Donnell and Mc. Geeney, Comparison of Saccharogenic and Phadebas.RTM. methods for amylase assay in biological fluids, Enzyme 18, 348 (1974), and Robyt and Whelan in Starch and its derivatives, 4th Ed. J. A. Radley, Ed. Chapman and Hall, London 1968 pp. 431-433.
Although these methods for determining .alpha.-amylase activity have proved to be valuable aids in discovering .alpha.-amylase levels in body fluids, they have certain limitations. In particular, these methods do not discriminate between the pancreatic and salivary types of .alpha.-amylase. Thus, only the total .alpha.-amylase activity, i.e. the sum of the salivary .alpha.-amylase activity and the pancreatic .alpha.-amylase activity, can be determined by these methods.
It would in many diagnostic situations be highly desirable not only to determine the total .alpha.-amylase concentration in body fluids, but also to separate between the two components thereof, i.e. salivary .alpha.-amylase and pancreatic .alpha.-amylase. In this manner it would be possible to discover hyper- or hypofunctioning states of pancreas or salivery glands, reflecting pathologic conditions such as pancreatitis (increased pancreatic .alpha.-amylase activity), ruptured ectopic pregnancy (increased salivary .alpha.-amylase activity), and others. It may also be mentioned, that a seemingly "normal" total .alpha.-amylase content of a body fluid may be composed of an abnormally high amount of one of the two enzymes and an abnormally low amount of the other.
Thus, there is a great need for rapid and reliable diagnostic methods, making it possible to determine the pancratic as well as the salivary .alpha.-amylase activity in body fluids. However, the gross similarity between these two enzymes has caused difficulties in measuring either of them in the presence of the other, and immunological methods of distinguishing between them have been unsuccessful. Pancreatic and salivary .alpha.-amylase can be separated by electrophoretic, electrofocusing or chromatographic methods [see E. G. S. Norby, Electrophoretic non-identity of human salivary and pancreatic amylases, Exp. Cell Res. 36; 663 (1964); Takeuchi, Matsushima and Sugimura, Separation of human .alpha.-amylase isoenzymes by electrofocusing and their immunologic properties, Clin. Chim. Acta 60, 207 (1975); Beck and Fridhandler, Clinical application of amylase isoenzyme analysis, Am. J. Gastroenterol 63, 457 (1975)]. However, these methods have the disadvantage of being tedious and time consuming.